Candida albicans is the most common fungal pathogen of AIDS patients. The infection most often involves the mucosal surfaces of the oral cavity, esophagus or vagina. In spite of the advent of triazoles as anti-fungals, management of AIDS patients with candidiasis is complicated by the development of drug-resistant C. albicans and non-albicans species. The spectrum of suitable alternatives is limited underlying the need to identify new targets, which can be exploited in the development of anti-fungals. Our approach has been to target non-mammalian phosphoproteins that are components of important signal transduction pathways of C albicans. To this end, we have identified a 2-component, histidine kinase (CaHKI) and response regulator genes (CaSSKl), both of which are essential for virulence of the organism, since knock-out strains in each gene were avirulent in comparison to parental, heterozygote and gene-reconstituted strains. The focus of this proposal will be on CaSSKl, the first response regulator isolated from C. albicans. Our reasons for emphasizing this gene are 2-fold. First, its absence results in avirulence. Second, a homologue of this gene in Schizosaccharomyces pombe provides essential adaptation functions for a number of stress conditions, including oxidative, high temperature and protein inhibitor stress. There are three specific aims in this proposal. In specific aim 1, in vitro studies will address oxidative stress. Expression will be measured by Northern analysis and competitive PCR in parallel with post-translational studies (phosphorylation). In specific aim 2, the virulence of CaSSKl-deleted strains (and their single-copy mates) will be evaluated in vaginal and oral models of candidiasis. Specific aim 3 will focus upon the identification of down-stream effectors of Casskip using subtractive hybridization and the yeast 2-hybrid system. The long-term objectives of this research are to expand the number of anti-C. albicans targets and information which will lead to the development of high-throughout assays for evaluating potential inhibitors.